One of the principle obstacles to the development of generic equivalents to biological and/or biotechnology-derived products is the absence of a practical, non-clinical method to compare the immunogenic potential of a proposed generic product with the immunogenic potential of an approved brand name reference product.
The immunogenic potential of a product is primarily based upon the three-dimensional shape and surface properties of the various molecular species present within that product. Immunogenicity can be further influenced by biological interactions between one or more species of the product and one or more endogenous compounds in a subject ingesting the product. Therefore, products that share the identical amino acid sequence can have disparate immunogenic potentials due to the presence of distinct molecular species within each product or due to the immunogenic properties of distinct formulation components of each product. Thus, while two proteins can share identical amino acid sequences (primary structure), the distinct molecular species of the proteins can comprise multiple three-dimensional conformations with distinct immunogenic properties. These differences in the three-dimensional structures of protein products can lead to differences in the immunogenic potential of the two protein products that share an identical primary amino acid sequence. Additionally, many of the biological products currently available (including both naturally-derived and biotechnologically-derived products) are not pure homogenous compounds. Rather, the products are heterogeneous mixtures of closely related molecular species. Even the products that are “substantially” homogeneous often contain low levels of impurities, such as degradation products, that are closely related to the “primary” product. In either case, the heterogeneous mixture or the presence of low level of impurities can contribute to the immunogenic potential of a given product. Thus, analytical methods which attempt to quantify immunogenicity by comparing only the amino acid sequence of two protein-base products and not the multiple three-dimensional molecular structures of the heterogeneous mixture are incapable of detecting the immunogenicity of the product resulting from the conformational differences between the distinct molecular species present within that product. Thus, there is a need in the art for methods which are capable of determining the distinct immunogenic potential profiles of protein-based products which have different formulations or are manufactured by distinct processes.
Methods of determining the immunogenic potential of biotechnological products, as well as other proteinaceous products, have been described, i.e., animal models have been used to assess the immunogenic potential of products. One such method includes the repeated administration of the protein of interest and subsequent evaluation of the animal for clinical signs. This method is not particularly useful because it is expected that all peptides larger than 5 kD will elicit an immune response in a non-homologous species. Therefore, the presence of an immune response is to be expected in such a protocol. Additionally, the mere quantification of these antibodies is not particularly insightful, since comparisons among different peptides and different species are not particularly informative. A second method for determining immunogenicity of a peptide product involves the repeated administration of the “final” product and subsequent animal evaluation. Such evaluations include observation of the animal for anaphylactic reactions and the measurement of immune complexes in the immunized animal. However, such methods are considered to be an inadequate means of determining the safety of the product in humans because of the lack of a direct correlation between the immune response that can occur in an animal model and the immune response that can occur in a human when exposed to the same potentially immunogenic product. Thus, human clinical trials represent the current benchmark for evaluating the immunogenic potential of biologically-derived pharmaceutical products.
In such human clinical trials, a biologically-derived pharmaceutical product is administered to a patient group and the group is evaluated for adverse immunological responses. Such adverse immunological responses include anaphylaxis, as well as the development of autoimmunity toward both the biologically-derived pharmaceutical product and the equivalent endogenous compound. For example, if a pharmaceutical preparation comprising recombinant DNA-produced insulin was administered to a human patient group and one of the patients were to develop anti-insulin antibodies in response to the recombinant protein, such anti-insulin antibodies could bind and inactivate any circulating insulin in that patient, including endogenously produced insulin. As a result, it would be very difficult to maintain adequate levels of insulin in said patient, even with the exogenous administration of insulin. Thus, human clinical trials which evaluate the immunogenicity of biotechnologically-derived pharmaceutical products can expose patient groups to an unnecessarily high degree of risk. Moreover, conducting human clinical trials is both time consuming and prohibitively costly. Further, the trials often provide spurious or variable results. Additionally, in the case of developing a generic equivalent to a brand-name biotechnologically-derived product, such human trials may preclude the filing of an abbreviate new drug application (ANDA), under section 505(j)(2)(C) of the Federal Food, Drug and Cosmetic Act (FD&C Act), thereby precluding the possibility of an 180 day market exclusivity period for the generic product under the Hatch-Waxman Act.
Thus, there exists a need in the art for a precise, non-clinical method of comparing the immunogenic profile of a test product with the immunogenic profile of a known reference product to evaluate the safety and efficacy of the test product relative to the safety and efficacy of the reference product.